The Polycomb Group Protein Bmi-1 Is Essential for the Growth of Multiple Myeloma Cells

Zainab Jagani,Christoph Lengauer,John Monahan,William R Sellers,Marion Dorsch,Rebecca Mosher,Paul Fordjour,Dmitri Wiederschain,Alice Loo,Dan He,Yung-Mae Yao,Markus Warmuth,Michael Morrissey

doi:10.1158/0008-5472.can-09-4229

Abstract

Bmi-1 is a member of the Polycomb group family of proteins that function in the epigenetic silencing of genes governing self-renewal, differentiation, and proliferation. Bmi-1 was first identified through its ability to accelerate c-Myc-induced lymphomagenesis. Subsequent studies have further supported an oncogenic role for Bmi-1 in several cancers including those of the breast, lung, prostate, and brain. Using a stable and inducible shRNA system to silence Bmi-1 gene expression, we show a novel role for Bmi-1 in regulating the growth and clonogenic capacity of multiple myeloma cells both in vitro and in vivo. Moreover, to elucidate novel gene targets controlled by Bmi-1, global transcriptional profiling studies were performed in the setting of induced loss of Bmi-1 function. We found that the expression of the proapoptotic gene Bim is negatively regulated by Bmi-1 and that Bim knockdown functionally rescues the apoptotic phenotype induced upon loss of Bmi-1. Therefore, these studies not only highlight Bmi-1 as a cancer-dependent factor in multiple myeloma, but also elucidate a novel antiapoptotic mechanism for Bmi-1 function involving the suppression of Bim.

Highlights

Bmi-1 was initially discovered through its ability to cooperate with c-Myc in the induction of lymphoma [1, 2] and was subsequently recognized as a member of the Polycomb group (PcG) family of proteins [3]
To ensure that such species represent Bmi-1, immunoblots were prepared in cell lines with and without Bmi-1 short hairpin RNA (shRNA)–mediated knockdown, and these protein species were each attenuated upon Bmi-1 knockdown (Supplementary Fig. S1C and E; Fig. 1B)
To determine whether Bmi-1 protein was required for MM cell growth, we infected RPMI-8226, KMS12BM, and LP1 cells with a robust inducible single-lentiviral vector knockdown reagent pLKO-Tet-On [28] containing either control nontargeting shRNA or two distinct Bmi-1–targeting shRNA sequences (Bmi-1 shRNA-1 or Bmi-1 shRNA-2), and polyclonal stable lines were obtained after puromycin selection

Summary

Introduction

Bmi-1 was initially discovered through its ability to cooperate with c-Myc in the induction of lymphoma [1, 2] and was subsequently recognized as a member of the Polycomb group (PcG) family of proteins [3]. The PcG proteins function within distinct multisubunit complexes and epigenetically regulate gene expression by altering chromatin states at specific promoters In several contexts, including lymphoma, mouse embryonic fibroblasts, and human fibroblasts, Bmi-1 exerts its oncogenic function, at least in part, by silencing the INK4a/ARF (Cdkn2a) tumor suppressor locus encoding the p16 (INK4a) and p19 (ADP ribosylation factor, ARF) proteins [5,6,7]. Several studies have shown that Bmi-1 is overexpressed in medulloblastomas [8], lung [9], breast [10], prostate cancers [11], and multiple myeloma (MM; ref. 12–14), among others.

Methods

Results

Conclusion